NMR clinical analyzers and related methods, systems, modules and computer program products for clinical evaluation of biosamples

ABSTRACT

Methods, computer program products and apparatus automate clinical NMR in vitro diagnostic analyzers. The clinical analyzer can automatically electronically monitor selected parameters and automatically electronically adjust parameters to maintain the analyzer within desired operational ranges. 
     The clinical NMR analyzers can be configured as a networked system with a plurality of clinical NMR analyzers located at different use sites; and at least one remote control system in communication with one or a plurality of clinical NMR analyzers, the at least one remote system configured to monitor selected local operating parameters associated with a respective clinical NMR analyzer.

RELATED APPLICATIONS

This application claims the benefit of priority of U.S. ProvisionalApplication Ser. No. 60/558,516, filed Apr. 1, 2004, the contents ofwhich are hereby incorporated by reference as if recited in full herein.

FIELD OF THE INVENTION

The present invention relates generally to NMR systems, and may beparticularly suitable for clinical NMR in vitro diagnostic systemscapable of analyzing biosamples.

BACKGROUND OF THE INVENTION

In the past, NMR detectors have been used to provide NMR LipoProfile®lipoprotein panel reports. The NMR detectors have been located in acentral testing facility with on-site support. The LipoProfile® report,available from LipoScience, Inc., located in Raleigh, N.C, is alipoprotein test panel that assesses a patient's risk of coronary arterydisease (“CAD”) and provides NMR-derived (quantitative analysis)lipoprotein measurement average low-density lipoprotein (LDL) particlesize as well as LDL particle number, the latter representing theconcentration or quantity (in concentration units such as nmol/L), andthe former representing the average size of the LDL particles (in nmunits) making up the LDL in the sample. HDL and VLDL subclassmeasurements can also be provided. See www.liposcience.com and U.S. Pat.No. 6,576,471 for exemplary reports of particular lipoprotein subclassparameters; the contents of the patent are hereby incorporated byreference as if recited in full herein.

It is known that NMR spectroscopic evaluation techniques can be used toconcurrently obtain and measure a plurality of different lipoproteinconstituents in an in vitro blood plasma or serum sample, as describedin U.S. Pat. No. 4,933,844, entitled Measurement of Blood LipoproteinConstituents by Analysis of Data Acquired from an NMR Spectrometer toOtvos and U.S. Pat. No. 5,343,389, entitled Method and Apparatus forMeasuring Classes and Subclasses of Lipoproteins, also to Otvos. Seealso, U.S. Pat. No. 6,617,167, entitled Method Of Determining PresenceAnd Concentration Of Lipoprotein X In Blood Plasma And Serum andco-pending co-assigned U.S. Provisional Patent Application Ser. No.60/513,795, entitled Methods, Systems and Computer Programs forAssessing CHD Risk Using Mathematical Models that Consider In VivoConcentration Gradients of LDL Particle Subclasses of Discrete Size. Thecontents of all the above patents and patent applications are herebyincorporated by reference as if recited in full herein.

As is well known to those of skill in the art, NMR detectors include anRF amplifier, an NMR probe that includes an RF excitation coil (such asa saddle or Helmholtz coil), a cryogenically cooled high-fieldsuperconducting magnet and an enclosed flow path that directs samples toflow serially, from the bottom of the magnet bore to a predeterminedanalysis location in the magnet bore. The NMR detector is typically ahigh-field magnet housed in a magnetically (and/or RF) shielded housingthat can reduce the magnetic field level that is generated to within arelatively small volume. NMR detectors are available from Varian, Inc.,having corporate headquarters in Palo Alto, Calif. and Bruker BioSpin,Corp., located in Billerica, Mass.

In operation, to evaluate the lipoproteins in a blood plasma and/orserum sample, the operator places the patient samples in a sample trayand an electronic reader correlates the sample to a patient, typicallyusing a bar code on the sample tray. The sample is aspirated from thesample container and directed to flow through the flow path extendingthrough the NMR detector. For detailed lipoprotein analysis, the NMRdetector may analyze the sample for 1-5 minutes to determine amplitudesof a plurality of NMR spectroscopy derived signals within a chemicalshift region of the proton NMR spectrum. These signals are derived bydeconvolution of the composite signal or spectrum and are compared topredetermined test criteria to evaluate a patient's risk of having ordeveloping coronary artery or heart disease.

In the past, a plurality of NMR spectrometers, all disposed at a centraltesting facility, have been used to carry out lipoprotein analysis onblood plasma samples to generate LipoProfile® test reports. The NMRspectrometers communicate with a local but remote computer (the computeris in a different room from the spectrometers) to allow the remotecomputer to obtain NMR spectra and analyze the NMR spectra to generatethe patient diagnostic reports with quantitative lipoprotein values.Unfortunately, an operator manually carries out adjustments to theequipment using a manually input quality control sample to adjust theline width. In addition, the sample handler does not communicate withthe NMR spectrometer and is not capable of electronically notifying thesystem of handling problems. The NMR spectrometer systems are complexand typically require dedicated on-site experienced operationaloversight.

In view of the above, there remains a need for improved NMR analyzersthat may be used in high-volume quantitative clinical applications atone or more remote locations.

SUMMARY OF EMBODIMENTS OF THE INVENTION

Certain embodiments of the present invention are directed at providingautomated NMR clinical analyzers that can be used without requiringdedicated on-site NMR support staff and/or undue technician support toreliably operate the NMR analyzers. The automated NMR clinical analyzerscan be configured to obtain quantitative analysis measurements that canbe used for in vitro diagnostics. In some embodiments, the automated NMRanalyzers can be configured to meet governmental medical regulatoryrequirements such as those described in applicable federal regulations,including those in 21 CFR (such as 21 CFR 820 and 21 CFR 11) for medicaldevices.

In some embodiments, the NMR analyzers can monitor and adjust selectedoperating parameters “on the fly” reducing the need for manualassistance and providing automated operation. The NMR analyzers caninclude interactive sample handlers that communicate with the NMRspectrometer and/or remote control system. The NMR clinical analyzerscan be configured to reliably run and obtain quantified clinicalmeasurements for diagnostic tests on high volume throughput ofbiosamples while reducing the amount of operator input or labor requiredto operate the automated analyzers. The NMR analyzers can be constructedand/or configured in such a manner as to be able to obtain PMA(pre-market approval) and/or 510(k) approval from the United States Foodand Drug Agency (“USFDA”) and/or corresponding foreign agencies.

Certain embodiments are directed to methods of operating a clinical NMRin vitro diagnostic analyzer. The methods include: (a) electronicallymonitoring data associated with selected equipment and/or environmentaloperational test conditions of a clinical NMR analyzer; (b)electronically determining whether the selected test conditions arewithin desired operational ranges based on the monitored data; (c)automatically adjusting operational parameters of selected components ofthe clinical NMR analyzer based on data obtained by the electronicallydetermining step; (d) obtaining NMR signal spectra of a biosample; and(e) analyzing the obtained NMR spectra to generate target clinicalmeasurements of the biosample.

Other embodiments are directed to clinical NMR in vitro diagnosticanalyzers. The analyzers include: (a) an automated sample handlerconfigured to automatically introduce biosamples into a magnet bore of ahigh-field superconducting magnet of a NMR spectroscopy instrumentassociated with a clinical NMR analyzer; (b) means for automaticallyobtaining NMR signal spectra of the biosamples; (c) means forautomatically electronically sensing data associated with selectedoperating parameters to verify that test conditions of the NMRdiagnostic analyzer are within target operating ranges; and (d) meansfor automatically electronically adjusting selected operating parametersbased on the verified test conditions.

Some embodiments are directed to clinical NMR in vitro diagnosticanalyzers that include: (a) an automated sample handler configured toautomatically introduce biosamples into a magnet bore of a high-fieldsuperconducting magnet of a NMR spectroscopy instrument associated witha clinical NMR analyzer; (b) a control circuit in electroniccommunication with the NMR spectroscopy instrument; and (c) a pluralityof electronic sensors disposed in the clinical NMR analyzer, theelectronic sensors in communication with the control circuit, theelectronic sensors configured to detect data associated with selectedoperating parameters to verify that selected conditions of the NMRdiagnostic analyzer are within target operating ranges. The clinical NMRanalyzer is configured to automatically electronically adjust selectedoperating parameters based on data provided by the electronic sensors sothat the clinical NMR analyzer operates within target process limits.

Still other embodiments are directed to computer program products forautomating clinical NMR in vitro diagnostic analyzers. The computerprogram products include a computer readable storage medium havingcomputer readable program code embodied in said medium. Thecomputer-readable program code includes: (a) computer readable programcode configured to automatically run an automated self-diagnosticquality control and/or calibration test for the clinical NMR in vitrodiagnostic analyzer; and (b) computer readable program code configuredto automatically electronically monitor selected operating parameters ofthe NMR in vitro diagnostic analyzer over time during operation.

Still other embodiments are directed to methods of analyzing undilutedplasma and/or serum by: (a) obtaining a proton NMR composite spectrum ofan undiluted biosample; and (b) generating a spectral deconvolution ofthe NMR composite spectrum using a predetermined doublet region of theproton NMR spectrum for spectral referencing and/or alignment.

The undiluted biosample may be neat serum and the doublet may comprise alactate doublet generally centered at about 1.3 ppm of the proton NMRspectrum. The undiluted sample can be serum that comprises glucose andthe doublet can be an anomeric proton signal from glucose in the serumthat is generally located at about 5.2 ppm of the NMR spectrum.

Another embodiment is directed to computer program products foranalyzing undiluted plasma and/or serum. The computer program productincludes a computer readable storage medium having computer readableprogram code embodied in the medium. The computer-readable program codecan include: (a) computer readable program code configured to obtain aproton NMR composite spectrum of an undiluted biosample, the proton NMRcomposite spectrum being devoid of a CaEDTA peak; and (b) computerreadable program code configured to generate a spectral deconvolution ofthe NMR composite spectrum using a predetermined doublet region of theproton NMR spectrum for spectral referencing and/or alignment.

Yet other embodiments are directed to clinical NMR in vitro analyzersthat include: (a) an automated sample handler for serially presentingrespective biosamples to an input port; (b) an enclosed flow pathconfigured to serially flow the respective biosamples presented by theautomated sample handler, wherein the enclosed flow path includes anon-magnetic rigid straight flow cell; (c) an NMR detector incommunication with an NMR flow probe, the NMR detector comprising ahigh-field cryogenically cooled superconducting magnet with a magnetbore, the flow probe configured to generally reside in the magnet bore,wherein the straight flow cell is configured and sized to extend intothe magnet bore and direct the samples to serially flow from a top ofthe magnet bore into the magnet bore during operation; and (d) aprocessor comprising computer program code for obtaining and analyzingNMR signal spectra of the biosamples to determine desired quantitativemeasurements of the respective biosamples.

Some embodiments of the present invention are directed to a networkedsystem of clinical NMR analyzers. The system includes: (a) a pluralityof clinical NMR analyzers located at different use sites; and (b) atleast one remote control (service/support) system in communication withthe plurality of clinical NMR analyzers. The at least one remote systemis configured to monitor selected local operating parameters associatedwith each clinical NMR analyzer.

In some embodiments, the remote system monitors the local NMR analyzersto inhibit down time and/or identify and correct process variablesbefore test data is corrupted to increase the reliability of theequipment and quantitative test results. The local and/or remote systemcan be configured to monitor predetermined process parameter data,service histories, cryogen use, patient test data, and the like.

In some embodiments, the local system can be configured to monitor andidentify process variation and generate an alarm that is sent to theremote system (local and/or remote site) for appropriate correctiveaction/investigation. In other embodiments, the remote system canmonitor the process variation and generate an alert to a service/supporttechnician at the remote site. Combinations of the local and remotemonitoring can also be used. The remote station can reduce the technicalsupport and/or operator knowledge needed at each local use site therebyallowing increased numbers of clinical analyzers to be used in fieldsites with relatively economic operational costs.

The local systems may generate and store an electronic history file ofselected operational parameters. The electronic history file can beconfigured to be accessed by the remote system. The local and/or remotesystem may be configured to automatically monitor process variables andstatistically analyze data corresponding to measurements of themonitored process variables to thereby perform an automated qualitycontrol analysis (such as maintain the parameters within a 3 sigmaand/or in some embodiments, a 6 sigma process limit). In someembodiments, the local system can be configured to automatically adjustoperating equipment to keep the process variables within a predeterminedstatistical variation responsive to the monitored data.

In some embodiments, the clinical NMR analyzers can be configured toautomatically adjust scaling of the NMR lineshape when the height and/orwidth thereof is outside a desired range. The local system can monitorRF excitation pulse power and automatically adjust the RF excitationpulse power if the power is outside a desired operating range and/orvaries from pulse to pulse by more than a predetermined amount and/orpercentage. In particular embodiments, the clinical NMR analyzers can beconfigured to disregard NMR signal data obtained when power variation ofthe RF pulses is greater than a predetermined amount.

Other embodiments are directed to methods of generating NMR-derivedquantitative measurement data for diagnostic clinical reports of patientbiosamples. The methods include: (a) automatically serially introducingbiosamples of interest into an NMR analyzer (which can be carried out byaspirating to an enclosed flow path that serially flows the biosamplesinto the NMR analyzer) having a NMR spectroscopy instrument with amagnet and a bore at a plurality of different clinical sites; (b)automatically correlating a patient identifier to a respective patientbiosample; (c) obtaining NMR derived quantitative measurements of thebiosamples for diagnostic reports; and (d) automatically monitoring theNMR analyzers at the different clinical sites from a remote monitoringstation.

In some embodiments the methods can include configuring theanalyzer/user to decide in situ how to analyze a particular patientbiosample. The obtaining step may include determining NMR derivedconcentration measurements of lipoproteins in an in vitro blood plasmaand/or serum sample.

In certain embodiments, the automated clinical NMR analyzer isconfigured with modular assemblies including: an automated samplehandling assembly; an NMR spectrometer; an NMR probe; and a sample flowpath to the NMR spectrometer each configured to releasably attach andoperate with its mating modular components thereby allowing ease ofrepair and/or field replacement.

The clinical NMR analyzers may be configured to automatically run anautomated self-diagnostic quality control test at startup. The analyzermay include computer program code that is configured to determine apatient's risk of having and/or developing CHD based on the NMR derivedquantitative measurements of the patient's respective biosample and/orcomputer program code that is configured to determine a patient's riskof having and/or developing Type II diabetes based on the NMR derivedquantitative measurements of the patient's respective biosample.

Yet additional embodiments are directed to clinical NMR in vitrodiagnostic apparatus for obtaining data regarding lipoproteinconstituents in a biosample. The apparatus includes: (a) an automatedsample handler system comprising a plurality of in vitro blood plasmaand/or serum samples; (b) an NMR spectrometer for serially acquiring anNMR composite spectrum of the in vitro blood plasma or serum sample incommunication with the automated handler system; (c) at least one sampleof validated control material configured to repeatedly controllably flowinto and out of the NMR spectrometer at desired times; and (d) aprocessor configured to receive data of the validated control material.The processor includes: (a) computer program code configured to definean a priori single basis set of spectra of validated reference controlmaterial; (b) computer program code configured to obtain NMR spectra ofthe validated control material; and (c) computer program code configuredto perform a spectral deconvolution of a CH₃ region of the obtained NMRspectra of the validated control material and comparing data associatedwith the spectral deconvolution of the CH₃ region with data associatedwith the a priori spectra of the validated control material to determinewhether the NMR analyzer is in a suitable operational status and/orready for diagnostic testing operation.

As will be appreciated by those of skill in the art in light of thepresent disclosure, embodiments of the present invention may includemethods, systems, apparatus and/or computer program products orcombinations thereof.

The foregoing and other objects and aspects of the present invention areexplained in detail in the specification set forth below.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a graph showing the chemical shift spectra of a representativesample of lipoprotein constituent subclasses.

FIG. 2 is a graph illustrating NMR spectra for a composite plasma sampleand the lipoprotein subclass and protein components thereof, with thepeaks for methyl groups being illustrated.

FIG. 3A is a schematic illustration of a single basis set of a prioridata used with the CH₃ region of a proton NMR spectra of a blood plasmaor serum sample according to embodiments of the present invention.

FIG. 3B is a graph of a proton NMR spectrum of serum with a lactatedoublet useable for spectral alignment according to embodiments of thepresent invention.

FIG. 3C is a graph of a proton NMR spectrum of serum with an anomericglucose doublet useable for spectral alignment according to embodimentsof the present invention.

FIG. 4 is a schematic illustration of a networked system of a pluralityof local clinical NMR analyzers that are in communication with anautomated remote service/support system according to embodiments of thepresent invention.

FIG. 5 is a schematic illustration of an in vitro diagnostic NMRanalyzer according to embodiments of the present invention.

FIG. 6 is a schematic illustration of an automated clinical NMR analyzeraccording to embodiments of the present invention.

FIG. 7 is a schematic illustration of another embodiment of an automatedclinical analyzer according to the present invention.

FIG. 8A is a schematic of NMR analyzer software architecture accordingto embodiments of the present invention.

FIG. 8B is a schematic of NMR analyzer software architecture accordingto embodiments of the present invention.

FIG. 9 is a flow chart of operations that can be carried out for an NMRanalyzer start-up and/or process evaluation procedure according toembodiments of the present invention.

FIG. 10 is a flow chart of operations that can be carried out to runcontrol samples of validated material through an NMR analyzer accordingto embodiments of the present invention.

FIG. 11 is a flow chart of normal-run mode or operation of an NMRanalyzer according to embodiments of the present invention.

FIG. 12 is a schematic diagram of a data processing system according toembodiments of the present invention.

DETAILED DESCRIPTION OF EMBODIMENTS OF THE INVENTION

The present invention will now be described more fully hereinafter, inwhich embodiments of the invention are shown. This invention may,however, be embodied in different forms and should not be construed aslimited to the embodiments set forth herein. Rather, these embodimentsare provided so that this disclosure will be thorough and complete, andwill fully convey the scope of the invention to those skilled in theart. In the drawings, like numbers refer to like elements throughout,and thickness, size and dimensions of some components, lines, orfeatures may be exaggerated for clarity. The order of operations and/orsteps illustrated in the figures or recited in the claims are notintended to be limited to the order presented unless stated otherwise.Broken lines in the figures, where used, indicate that the feature,operation or step so indicated is optional unless specifically statedotherwise.

It will be understood that when a feature, such as a layer, region orsubstrate, is referred to as being “on” another feature or element, itcan be directly on the other feature or element or intervening featuresand/or elements may also be present. In contrast, when an element isreferred to as being “directly on” another feature or element, there areno intervening elements present. It will also be understood that, when afeature or element is referred to as being “connected”, “attached” or“coupled” to another feature or element, it can be directly connected,attached or coupled to the other element or intervening elements may bepresent. In contrast, when a feature or element is referred to as being“directly connected”, “directly attached” or “directly coupled” toanother element, there are no intervening elements present. Althoughdescribed or shown with respect to one embodiment, the features sodescribed or shown can apply to other embodiments.

Unless otherwise defined, all terms (including technical and scientificterms) used herein have the same meaning as commonly understood by oneof ordinary skill in the art to which this invention belongs. It will befurther understood that terms, such as those defined in commonly useddictionaries, should be interpreted as having a meaning that isconsistent with their meaning in the context of the relevant art andthis application and should not be interpreted in an idealized or overlyformal sense unless expressly so defined herein. As used herein, theterm “and/or” includes any and all combinations of one or more of theassociated listed items.

As used herein, the singular forms “a”, “an” and “the” are intended toinclude the plural forms as well, unless the context clearly indicatesotherwise. It will be further understood that the terms “comprises”and/or “comprising,” when used in this specification, specify thepresence of stated features, integers, steps, operations, elements,and/or components, but do not preclude the presence or addition of oneor more other features, integers, steps, operations, elements,components, and/or groups thereof.

The term “biosample” includes whole blood, plasma, serum, urine,cerebral spinal fluid (CSF), lymph samples, stool samples, tissues,and/or body fluids in raw form and/or in preparations. The biosamplescan be from any target subject. Subjects', according to the presentinvention, can be any animal subject, and are preferably mammaliansubjects (e.g., humans, canines, felines, bovines, caprines, ovines,equines, rodents (mice, rats, hamsters, guinea pigs or others),porcines, primates, monkeys, and/or lagomorphs). The animals can belaboratory animals or non-laboratory animals, whether naturallyoccurring, genetically engineered or modified, and/o whether beinglaboratory altered, lifestyle and/or diet altered or drug treated animalvariations.

The term “clinical” with respect to data measurements means qualitativeand/or quantitative measurements that can be used for therapeutic ordiagnostic purposes, and typically for diagnostic purposes and meets theappropriate regulatory guidelines for accuracy, depending on thejurisdiction or test being performed. The term “clinical” with respectto NMR analyzer is described above in the Summary section of thespecification.

The term “automatic” means that substantially all or all of theoperations so described can be carried out without requiring activemanual input of a human operator, and typically means that theoperation(s) can be programmatically directed and/or carried out. Theterm “electronic” means that the system, operation or device cancommunicate using any suitable electronic media and typically employsprogrammatically controlling the communication between a control systemthat may be remote and one or more local NMR analyzers using a computernetwork.

The term “computer network” includes one or more local area networks(LAN), wide area networks (WAN) and may, in certain embodiments, includea private intranet and/or the public Internet (also known as the WorldWide Web or “the web”). The term “networked” system means that one or aplurality of local analyzers can communicate with at least one remote(local and/or offsite) control system. The remote control system may beheld in a local “clean” room that is separate from the NMR clinicalanalyzer and not subject to the same biohazard controlrequirements/concerns as the NMR clinical analyzer.

As will be appreciated by one of skill in the art, the present inventionmay be embodied as an apparatus, a method, a data or signal processingsystem, and/or a computer program product. Accordingly, the presentinvention may take the form of an entirely software embodiment, or anembodiment combining software and hardware aspects. Furthermore, certainembodiments of the present invention may take the form of a computerprogram product on a computer-usable storage medium havingcomputer-usable program code means embodied in the medium. Any suitablecomputer readable medium may be utilized including hard disks, CD-ROMs,optical storage devices, or magnetic storage devices.

The computer-usable or computer-readable medium may be, but is notlimited to, an electronic, magnetic, optical, superconducting magnetic,infrared, or semiconductor system, apparatus, device, or propagationmedium. More specific examples (a nonexhaustive list) of thecomputer-readable medium would include the following: an electricalconnection having one or more wires, a portable computer diskette, arandom access memory (RAM), a read-only memory (ROM), an erasableprogrammable read-only memory (EPROM or Flash memory), an optical fiber,and a portable compact disc read-only memory (CD-ROM). Note that thecomputer-usable or computer-readable medium could even be paper oranother suitable medium, upon which the program is printed, as theprogram can be electronically captured, via, for instance, opticalscanning of the paper or other medium, then compiled, interpreted orotherwise processed in a suitable manner if necessary, and then storedin a computer memory.

Computer program code for carrying out operations of the presentinvention may be written in an object oriented programming language suchas Java®, Smalltalk, Python, Labview, C++, or VisualBasic. However, thecomputer program code for carrying out operations of the presentinvention may also be written in conventional procedural programminglanguages, such as the “C” programming language or even assemblylanguage. The program code may execute entirely on the user's computer,partly on the user's computer, as a stand-alone software package, partlyon the user's computer and partly on a remote computer or entirely onthe remote computer. In the latter scenario, the remote computer may beconnected to the user's computer through a local area network (LAN) or awide area network (WAN), or the connection may be made to an externalcomputer (for example, through the Internet using an Internet ServiceProvider).

The flowcharts and block diagrams of certain of the figures hereinillustrate the architecture, functionality, and operation of possibleimplementations of analysis models and evaluation systems and/orprograms according to the present invention. In this regard, each blockin the flow charts or block diagrams represents a module, segment,operation, or portion of code, which comprises one or more executableinstructions for implementing the specified logical function(s). Itshould also be noted that in some alternative implementations, thefunctions noted in the blocks may occur out of the order noted in thefigures. For example, two blocks shown in succession may in fact beexecuted substantially concurrently or the blocks may sometimes beexecuted in the reverse order, depending upon the functionalityinvolved.

Embodiments of the present invention may be used to analyze any in vitrobiosample. The biosample may be in liquid, solid, and/or semi-solidform. The biosample may include tissue, blood, biofluids, biosolids andthe like. However, as noted above, the automated clinical NMR analyzermay be particularly suitable to analyze lipoprotein data in in vitroblood serum and/or plasma samples. The small person-to-person variationsin the lineshapes of the lipoprotein classes are caused by the subclassheterogeneity known to exist within each of these lipoprotein classes.FIG. 1 shows the lineshapes and chemical shifts (positions) for a numberof subclasses of lipoproteins. As shown in FIG. 1, the chemical shiftsand lineshape differences between the different subclasses are muchsmaller than those between the major lipoprotein classes, but arecompletely reproducible. Thus, differences among the NMR signals fromthe plasma of individuals are caused by differences in the amplitudes ofthe lipid resonances from the subclasses present in the plasma, which inturn are proportional to their concentrations in the plasma. This isillustrated in FIG. 2, in which the NMR chemical shift spectra of ablood plasma sample is shown. The spectral peak produced by methyl (CH₃)protons 60 (shown as a solid line) is shown for the blood serum samplein FIG. 2. The spectral peak 61 (shown as a dotted line) in FIG. 2 isproduced by the arithmetic sum of the NMR signals produced by thelipoprotein subclasses of the major classes VLDL, LDL, HDL, proteins andchylomicrons, as illustratively shown by certain of the subclasses inFIG. 1. It can be seen that the lineshape of the whole plasma spectrumis dependent on the relative amounts of the lipoprotein subclasses whoseamplitudes change (sometimes dramatically) with their relativeconcentrations in the plasma sample.

Since the observed CH₃ lineshapes of whole plasma samples are closelysimulated by the appropriately weighted sum of lipid signals of theirconstituent lipoprotein classes, it is possible to extract theconcentrations of these constituents present in any sample. This isaccomplished by calculating the weighting factors which give the bestfit between observed blood plasma NMR spectra and the calculated bloodplasma spectra. Generally speaking, the process of NMR lipoproteinanalysis can be carried out by the following steps: (1) acquisition ofan NMR “reference” spectrum for each of the “pure” individualconstituent lipoprotein classes and/or subclasses of plasma or serum ofinterest and/or related groupings thereof; (2) acquisition of a wholeplasma or serum NMR spectrum for a sample using measurement conditionssubstantially identical to those used to obtain the reference spectra;and (3) computer deconvolution of the NMR spectrum in terms of theconstituent classes and/or subclasses (or related groupings thereof) togive the concentration of each lipoprotein constituent expressed as amultiple of the concentration of the corresponding lipoproteinreference.

Although the procedure can be carried out on lipoprotein classes,carrying out the process for subclasses of lipoproteins can decrease theerror between the calculated lineshape and the NMR lineshape, thusincreasing the accuracy of the measurement while allowing forsimultaneous determination of the subclass profile of each class.Because the differences in subclass lineshapes and chemical shifts aresmall, for certain applications, it may be important to correctly alignthe reference spectrum of each subclass with the plasma spectrum.

The alignment of these spectra can be accomplished by the alignment ofcontrol peaks in the spectra, which are known to respond in the samemanner to environmental variables, such as temperature and samplecomposition, as do the lipoprotein spectra. As is known, one suchsuitable alignment peak is the peak produced by CaEDTA found in prepared(diluted) sample mixtures, although other EDTA peaks or suitable peakmay be utilized. By alignment of the spectra, the small variations inthe subclasses' lineshapes and chemical shifts may be exploited toproduce higher accuracy and subclass profiles.

Further description of these methods can be found in U.S. Pat. Nos.4,933,844 and 5,343,389 to Otvos. The mathematics used in the lineshapefitting process (i.e., least squares fit of an unknown function in termsof a weighted sum of known functions) is well known and is described inmany textbooks of numerical analysis, such as F. B. Hildebrand,Introduction to Numerical Analysis, 2nd edition, pp. 314-326, 539-567,McGraw-Hill, 1975.

Validation Control Material and Operational Status Evaluation

In the past, as part of start-up or periodic quality assessment, atleast two types/levels of control material samples were introduced intothe NMR spectrometer and multiple NMR derived lipoprotein parameterswere assessed (compared to stored values) for conformance to expectedresults for quality control review.

In some embodiments of the present invention, it is contemplated thatthe multiple variables previously reviewed can be reduced to a singlevariable by performing spectral deconvolution of the CH₃ region of thespectra or other suitable region for at least one validation controlmaterial sample. The analyte NMR lineshape can be deconvoluted usingmultivariate analysis with non-negative constraints. See, e.g., Lawson,C. L., Hanson, R. J. Solving Least Squares problems. Philadelphia, Pa.:SIAM, 1995, pp. 160-165.

The analyte spectra array consists of “m” discrete data points denotedP_(i) ⁰, where i=1, 2 . . . m. The method for fitting the validatedcontrol spectrum, P_(i) ⁰, with a linear combination of n constituentspectra is based on the premise that there are a set of coefficients,c_(j), corresponding to the contributions of component j, and acoefficient, c_(p) ¹, corresponding to the imaginary portion of thesample plasma spectrum, such that for each data point, P_(i) ⁰≈P_(i)^(c), where

$\begin{matrix}{P_{i}^{c} = {{\sum\limits_{j = 1}^{n}{c_{j}V_{ji}}} + {c_{p}^{I}{V_{i}^{I}.}}}} & {{EQUATION}\mspace{14mu}(1)}\end{matrix}$

The best fit can be obtained by minimizing the root mean square error ina manner analogous to that previously described in U.S. Pat. No.6,617,167, except that V_(j) represents only the single (j=1) basis setof the validated control spectra array stored in the computer. Thecontents of this patent are hereby incorporated by reference as ifrecited in full herein.

The correlation coefficient, r, of the fit of control spectra of the CH₃region as a function of the stored validated control spectra will beused along with coefficient c_(j) to determine the acceptability ofstatus of the analyzer to acquire clinical data. In certain embodiments,both r and c_(j) can be as chosen to be as close to 1.0 as practicableand/or possible. Acceptable limits for deviation from 1.0 can beestablished in consonant with standard clinical practices mandated byCLIA.

The phrase “validation control material sample” refers to a priori orknown measurement values of a known reference sample, the known samplecorresponding to those types of samples that will be undergoingevaluation using the equipment and analysis software (whatever biotype,i.e., blood plasma or serum, urine, etc). The spectral deconvolution ofthe CH₃ region of the spectra of the control material can be carried outusing the single basis set of the stored spectra of the validated knowncontrol samples. Thus, a known validation sample can be analyzed and itsassociated values can be stored as known or control values.Periodically, the validation control sample can be reanalyzed by the NMRsystem to confirm that the test values conform to the stored (expectedvalues). The NMR analyzer can be configured to flag or alert when thereis undue departure from predetermined norms so that the system can berecalibrated.

The validation control sample and validation control protocol cantypically be run at start-up (each shift or daily) and at increasedintervals as needed. The increased intervals may be based on signaldegradation of the proton NMR spectrum lineshape (width/height), when anunknown sample is quantified outside normal bounds and/or upon otherautomatically detected and monitored parameters.

Undiluted Samples

In certain embodiments, the NMR clinical analyzers 10 (FIG. 4) can beconfigured to analyze undiluted (neat) plasma and/or serum.Unfortunately, a CaEDTA peak may not appear when the sample is undilutedserum, which can impede spectral referencing for deconvolution. Thus, incertain embodiments, as shown in FIG. 3B, a lactate doublet 66 generallycentered about 1.3 ppm in the proton NMR spectrum of serum can be usedfor spectral referencing and alignment for NMR derived quantificationanalysis (such as lipoprotein quantification of serum samples). In otherembodiments, as shown in FIG. 3C, an anomeric proton signal from theglucose in serum can appear as a doublet 67 at about 5.2 ppm and thisdoublet may also be used (with or alone) as an anchor point for spectralalignment.

Network of Clinical Analyzers

As shown in FIG. 4, certain embodiments of the invention are directed toa networked system 18 of clinical NMR analyzers 10. The networked system18 includes at least one clinical NMR analyzer 10 that communicates withat least one remote system 15. Typically, a plurality of clinical NMRanalyzers 10 located at a common local use site communicate with arespective at least one remote service/support system 15. The at leastone remote system 15 can be configured to monitor selected localoperating parameters associated with each clinical NMR analyzer 10. Insome embodiments, each local site may include a plurality (at least two)NMR analyzers 10, which may be configured to communicate with each otherand/or at least one remote control system 15. The at least one remotecontrol system 15 may be configured as a common local or offsite controlstation for a plurality of different local analyzers 10 (typically forall of the local analyzers at a use site). The at least one remotecontrol system 15 can be a plurality of generally independent stationsconfigured to communicate with one or selected local analyzers 10. Inother embodiments, the at least one control system 15 can be a pluralityof remote control systems 15 that may be in communication with anotheroffsite control station 15′ as optionally shown in FIG. 4. Eachrespective local analyzer 10 can communicate with a common remotecontrol system 15 or a plurality may communicate with different controlsystems and/or sites. The local analyzers 10 may also be configured tooperate independently of the others and/or not to communicate with eachother. The broken line box 15R drawn around the remote control box inFIG. 4 illustrates that the remote control system(s) 15 can be locatedon-site (in the same facility) but in a room 15R that is enclosed andaway from the NMR Analyzers 10 so as to not be under the same biohazard,laboratory access/cleanliness or operation restrictions as the NMRclinical analyzer itself 10.

In some embodiments some of the local analyzers 10 may be configured tocommunicate with each other directly or indirectly using the controlsystem 15, such as, but not limited to, those at affiliated locations ora common local site. The communication can be electronic communicationsuch as (a) wireless, which may be carried out using mobilecommunications and/or satellite systems, (b) via an intranet, (c) via aglobal computer network such as the Internet, and/or (d) use a POTS(land based “plain old telephone system”). The system 18 may usecombinations of communications systems.

The local analyzers 10 may be controlled by the remote system 15 in amanner that allows for interactive adjustment during operation, such asduring the NMR analysis and/or start-up or calibration mode. As such,the operational and/or test analysis data can be relayed to the remotecontrol system 15 in substantially “real-time”. The NMR analyzers 10 canbe configured to interactively communicate with the remote controlsystem 15 to allow “smart” monitoring status. For example, the NMRanalyzer 10 can automatically send a signal alerting the control system15 when a test is complete for a subject, allowing the control system 15to timely obtain the data therefrom and generate the test report usingthe data.

In some particular embodiments, the system 18 can include a dataprocessing system, which comprises a web server. In particular, the dataprocessing system may be an Internet Appliance, such as a PICOSERVER®appliance by Lightner Engineering located in San Diego, Calif. (see alsowww.picoweb.net) or other such web servers, including, but not limitedto, those available from Axis Communications, or PICOWEB, RABBIT, andthe like. The data processing system can receive commands from thesupport site 15 and controls certain operational parameters of thesystem 10. The data processing system can also include a TCP stack andEthernet NIC to provide the communication link between the computernetwork 10 and the test administration site 15.

The processing system can provide information about the local analyzers10 to the administration site 15 as web pages which may be predefinedand stored at the local device 10. Such web pages may also bedynamically generated to incorporate test specific information. The webpages may be Hypertext Markup Language (HTML) common gateway interface(CGI) web pages which allow for user input by a client, such as a webbrowser, of a user at the test administration site 15. The web pages mayalso be or include Java scripts, Java applets or the like which mayexecute at the test administration site so as to control operations ofan administration data processing system at the administration site 15.As will be appreciated by those of skill in the art, other mechanismsfor communicating between a web server and a client may also beutilized. For example, other markup languages, such as Wireless MarkupLanguage (WML) or the like, for communicating between the local device10 and the administration site 15 may be utilized.

In certain embodiments, operations of a web server and a web client caninclude a web browser as the administration site 15 that requests aninitial web page from the web server of the local device 10. Such arequest may take the form of a Hypertext Transfer Protocol (HTTP)request to the IP address of the web server of the local device. The IPaddress may be pre-assigned to the local device 10 or may be dynamicallyassigned when the local device 10 attaches to the network 15. Thus, theweb browser may know in advance the IP address of the local devices 10or may be notified of the IP address as part of a setup procedure.

When the local device 10 receives the request for the initial web page,it sends the initial web page and a Java applet which causes the webbrowser to periodically reload its current web page. Alternatively,“push” technology could be employed by the server to push data to theweb browser when status is to be updated. The rate at which the web pageis reloaded may be based on the type of data relayed or detected and/orthe web page being displayed. Similarly, the rate may also be based onthe type of network connection utilized such that for slower connectiontypes the refresh rate could be reduced. In some embodiments, the Javaapplet could be generated once with the initial web page, while inothers the Java applet could be provided with each web page, and therefresh rate could be based on the particular web page provided. Forexample, a setup web page could be refreshed less often then a teststatus web page (or not at all).

In any event, after the initial web page is provided to the web browser,the web server of the local devices 10 waits for a subsequent requestfor a web page. When a request is received, it may be determined if therequest is for a response to an operational status inquiry, such aslineshape width and/or height of the last two samples, which is to beincluded in the responsive web page. If so, then the web page may berevised to indicate the information. In any event, it may also bedetermined if the request specifies parameters for the inquiry by, forexample, providing a CGI request which reflects user input to the webbrowser. If so, the parameters are set based on the CGI specificationsand the web page corresponding to the URL of the request is returned tothe web browser. If the inquiry is terminated, then operations mayterminate. Otherwise, the web server waits for the next request from theweb browser.

In some embodiments, the clinical NMR analyzers 10 include a high fieldNMR superconducting magnet and the remote system automatically obtainsdata regarding homogeneity of the magnetic field generated by thesuperconducting magnet. The homogeneity data can include data regardingthe lineshape characteristics of biosamples undergoing analysis (whichcan indicate a degradation in homogeneity over time). In someembodiments, the local NMR analyzer 10 generates and stores anelectronic history file of selected operational parameters. The localNMR analyzers 10 can be configured to review and generate an automaticapproval of each sample test results and/or a retest (reject) decision.

The history file can be configured to be electronically accessible bythe remote system 15. In some embodiments, the local analyzers 10 and/orremote system 15 are configured to automatically monitor processvariables and statistically analyze data corresponding to measurementsof the monitored process variables to thereby perform an automatedquality control analysis. In particular embodiments, the local systems10 are configured to automatically adjust operating equipment to keepthe process variables within a predetermined statistical variationresponsive to the monitored data. The local systems can be configured toautomatically generate an alert when an abnormal operating condition isdetected. In other embodiments, the remote system 15 is configured toautomatically generate an alert when an abnormal operating condition isdetected at the local NMR analyzer site(s) 10. The local analyzers 10can be configured to generate an electronic service log and/or anelectronic process history log that is electronically accessible by theremote system 15.

The local analyzers 10 can be configured to automatically detecttemporally relevant data of selected operational parameters at desiredintervals and generate an electronic maintenance file thereof, and thelocal NMR analyzers 10 can be configured to electronically store theirrespective maintenance files for electronic interrogation by the remotesystem 15. The selected operational parameters can include the NMRsignal lineshape and/or scaling thereof of one or more patient samples.The maintenance file may include respective patient sample identifierscorrelated to selected operational parameters measured at a time the NMRsignal of the patient sample was obtained, and may also include a timeand/or date stamp or data. The local NMR analyzers 10 can be configuredto generate an electronic maintenance file of selected operationalparameters for each sample processed. The local NMR analyzers 10 canelectronically store (at least temporarily) sample data correlated to anaccession patient identifier and/or sample dilution factor.

In some embodiments, the local analyzer 10 can generate an electroniclog of NMR sample data that is analyzed for one or more biosamples andthe log can be configured to be accessible by the remote system 15. Incertain embodiments, an operator or service program at the remote system15 determines when to send (and places the service order for) technicalsupport onsite to the local clinical analyzers 10.

In particular embodiments, the remote system 15 automatically controlsselected features of the local clinical NMR analyzers 10. The local NMRanalyzers 10 can be configured with a user interface that accepts localuser input to select a report format and/or sample variables of interestfor NMR analysis, thereby allowing customizable report formats bysite/region. Patient reports generated by the analyzers 10 at each localclinical NMR analyzer site can have a site identifier thereon and thereport can be generated in electronic and/or paper form.

To help monitor the number of tests performed, the remote system 15 canautomatically obtain data regarding the number of patient samplesanalyzed over a desired interval on each clinical NMR analyzer 10. Thismonitoring can allow the remote control system to order consumablesbased on projected and/or actual needs customized to a particular site.

In some embodiments, each NMR analyzer 10 can include an electroniclibrary of predetermined computer program functions that refer to a NMRnormalization factor used to carry out quantification measurements. TheNMR analyzers 10 can be configured to obtain NMR derived concentrationmeasurements of lipoproteins in a blood plasma and/or serum sample. Insome embodiments, the NMR analyzers 10 can be configured to obtain NMRderived concentration measurements of one or more of LDL, HDL, and/orVLDL subclass particles in a blood plasma and/or serum sample and/orconfigured to determine: (a) a patient's risk of having and/ordeveloping CHD based on the lipoprotein measurements; and/or (b) apatient's risk of having and/or developing Type II diabetes or otherinsulin resistance disorders.

In certain embodiments, each clinical NMR analyzer 10 can be configuredto automatically execute a start-up self-diagnostic and/ortuning/calibration routine and relay abnormal data regarding same to theremote control system 15. The clinical NMR analyzers 10 can beconfigured to automatically monitor the NMR signal lineshapes, and/ordetermine a height and/or width thereof, over time, to monitor ifadjustments to equipment are indicated. For example, the clinical NMRanalyzers 10 include a high field superconducting magnet and theclinical NMR analyzers can be configured to automatically shim the NMRspectroscopic magnetic field to provide increased homogeneity if theline widths degrade beyond a desired amount.

In some embodiments, the clinical NMR analyzers 10 can be configured toautomatically adjust scaling of the NMR lineshape of the proton NMRspectrum of the biosample when the height and/or width thereof isoutside a desired range.

In some embodiments, one of the selected operational parametersmonitored for can be RF excitation pulse power. The clinical NMRanalyzers 10 can be configured to automatically adjust the RF excitationpulse power (increase or decrease the RF amplifier, if the power isoutside a desired operating range and/or varies from pulse to pulse(and/or sample to sample) by more than a predetermined amount and/orpercentage. The clinical NMR analyzers 10 can be configured to disregardand/or invalidate NMR signal data obtained when power variation of theRF pulses is greater than a predetermined amount. In some embodiments,when large RF power changes are detected, the analyzer 10 can beconfigured to disregard, flag as error-prone and/or invalidate thesample data. In some embodiments, increased accurate control of RF powermonitoring can be obtained by using a controlled sample introduced intothe analyzer 10 at desired intervals, such as a standard solutioncontaining TMA.

The networked system 18 can be configured to monitor, in substantiallyreal time, at least intermittently and/or at desired intervals, certainparameters associated with the operational status of the NMR analyzer 10during operation. The system 18 may go into a standby mode duringnon-active periods (down shifts), but monitor for certain majorparameters, such as cryogen level, electronic circuitryover-temperature, and the like.

Automated Clinical NMR Analyzer

FIG. 5 is a schematic diagram of one example of an in vitro diagnostic(IVD) clinical NMR analyzer 10. As shown, the analyzer 10 includes anNMR detector 50, an enclosed flow path 65, an automated sample handler70, and a controller/processor 80 (shown as a CPU) with operationalsoftware 80 s. The term “NMR detector” may also be known as an NMRspectrometer as will be appreciated by those of skill in the art. TheNMR detector 50 includes a magnet, typically a cryogenically cooled highfield superconducting magnet 20, with a magnet bore 20 b, a flow probe30, and RF pulse generator 40. The term “high-field” magnet refers tomagnets that are greater than 1 Tesla, typically greater than 5 Tesla,and more typically greater than about 9 Tesla. Magnetic fields greaterthan about 13 Tesla may, in some situations, generate broaderlineshapes, which in some analysis of some biosamples, may not bedesirable. The flow probe 30 is in communication with the RF pulsegenerator 40 and includes an RF excite/receive circuit 30 c, such as aHelmholtz coil. However, as will be appreciated by those of skill in theart, other excite/receive circuit configurations may also be used.

It is noted that although illustrated as a system that serially flowsbiosamples using a flow cell 60, other sample handlers 70 and biosampleintroduction means can be used. For example, the biosample can beprocessed as it is held in a respective tube or other sample container(not shown). In some embodiments, each of the modular components of theNMR analyzer 10 may be sized and configured to fit within a singlehousing or enclosure.

Field homogeneity of the detector 50 can be adjusted by shimming on asample of about 99.8% D₂O until the spectral linewidth of the HDO NMRsignal is less than 0.6 Hz. The 90° RF excitation pulse width used forthe D₂O measurement is typically about 6-7 microseconds. Other shimmingtechniques can also be used. For example, the magnetic field can beautomatically adjusted based on the signal lineshape and/or a width orheight thereof. The NMR detector 50 may optionally include a gradientamplifier in communication with gradient coils 41 held in the magnetbore 20 b as is well known to those of skill in the art, and thegradient system may also be used to help shim the magnet.

During operation, the flow probe 30 is held inside the magnet bore 20 b.The flow probe 30 is configured to locate the flow probe RF circuitry 30c within the bore 20 b to within about +/−0.5 cm of a suitablyhomogeneous portion of the magnetic field B₀. The flow probe 30 is alsoconfigured to receive the flow cell 60 that forms part of the biosampleenclosed flow path 65. The flow cell 60 typically includes a largerholding portion 60 h that aligns with the RF circuitry 30 c of the flowprobe 30. The flow cell 60 is configured to remain in position with theholding portion 60 h in the magnet bore 20 b and serially flowbiosamples to the holding portion 60 h, with successive biosamples beingseparated by a fluid (typically air gaps) to inhibit cross-contaminationin a flowing stream. The samples may be introduced as a train ofsamples, but are more typically introduced (injected) one at a time. Thebiosample is typically held in the holding portion 60 h for betweenabout 1-5 minutes during which time a proton NMR spectrum is obtainedand electronically correlated to the sample accession number oridentifier (i.e., a patient identifier). The flow cell 60 can be formedof a non-magnetic material that does not degrade the performance of theNMR detector 50. Typically, the flow cell 60 is formed of a suitablegrade of silicate (glass) material, however, other magnetic-friendlynon-porous materials may be used including ceramics, polymers, and thelike.

A magnetically-friendly optic viewing scope (such as a fiber opticsystem) may be used to allow a user and/or the system 10 to visuallymonitor conditions in the magnet bore 20 b (i.e., position of the probe,leaks or the like) (not shown). The viewing scope can be mounted to thebore or made integral to the flow cell 60 or the flow probe 30.Similarly, at least one leak sensor can be placed to automaticallydetect fluid leakage, whether biosamples, cleansers or cryogens. If theformer, a leak sensor can be used to detect leaks caused by flow pathdisruption; if the latter a gas sniffer type sensor can be used. The gassensor can be located away from the probe. Cryogen level sensors canalso be used to monitor the level of the liquid (helium and/or nitrogen)to allow for automated supply orders, identification of an increased userate (which may indicate a magnet problem), and the like.

In the embodiment shown, the flow cell 60 is in fluid communication witha waste receptacle 61 at one end portion and a sample intake 73 on theother end portion. In certain embodiments, the analyzer 10 is configuredto flow the samples from top to bottom using a flow cell 60 that has amajor portion that is substantially straight (i.e., without bends) toreduce the length of the flow path 60 and/or to reduce the likelihoodthat the bends in a flow path will block the flow. In some embodiments,the flow cell 60 is entirely straight. In particular embodiments, theentire flow path 65 may be straight throughout its length (includingportions upstream and downstream of the flow cell 60, from intake todischarge into the waste container). In other embodiments, elastomeric,typically polymeric, conduit and/or tubing (which may comprise TEFLON)can be used to connect the flow cell 60 to the sample intake portion ofthe flow path 65 and the conduit and/or tubing may be bent to connect tomating components as desired. However, it the conduit/tubing extend intothe magnet bore 20 b, then that part of the flow path 65 may also beconfigured to be straight as discussed with respect to the flow cell 60.

In some embodiments, the flow cell 60 has an inner diameter of betweenabout 0.5 mm to about 0.8 mm upstream and downstream of the holdingportion 60 h. The downstream portion is typically at least about 0.8 mmto inhibit clogs in the flow system. The holding portion 60 h may have adiameter that is between about 1.0 mm- to about 4.0 mm.

The biosamples may be configured in appropriate sample volumes,typically, for blood plasma or serum, about 0.5 ml. For whole plasma, areduced sample size of about 50-300 microliters, typically about 60-200microliters, and more typically between about 60-100 microliters may bedesired. In some embodiments, the sample flow rate may be between about2-6 ml/min to flow the sample to the holding portion 60 h for the NMRdata collection and associated analysis.

Still referring to FIG. 5, the automated sample handler 70 may beconfigured to hold a plurality of samples 70 s in suitable samplecontainers 70 c and present the samples 70 s in their respectivecontainer 70 c to an intake member 72 that directs the sample into theenclosed flow path 65. The sample bed 71 may hold about 50-100 samplesin containers. In some embodiments, the bed 71 may optionally beconfigured to provide and/or held in a refrigerated or cooled enclosedcompartment. In other embodiments, conventional small and/or large racksof sample tubes can be used. Typically, the intake member 72 isconfigured to aspirate the sample into the flow path 65. As shown, theintake member 72 comprises a pipetter and/or needle that withdraws thedesired sample amount from the container 70 c, and then directs thesample (typically via injection through an injection port) into aconduit 73 that is in fluid communication with the flow cell 60. Thepipette may rotate about 180 degrees to access tray samples or a labautomation system (TLA, workcell, etc.). However, other sample transfermeans may also be used. In other embodiments, the intake member 72 canbe in direct communication with the flow cell 60 without the use of anintermediate conduit 73. In particular embodiments, the samples may bedirectly aspirated from a source tube on the sample handler tray. Thesample handler system 70 can be configured to provide rapid flowcleaning and sample delivery. In particular embodiments, the handlersystem 70 can be configured to operate on about a 1-minute or less cycle(excluding NMR data acquisition) while reducing dilution and/orcarryover.

A multi-port valve (which may replace or be used with the injectionport) may be used to help reduce unwanted sample dilution due to flowcleaning carried out between samples. In certain embodiments, the intakemember 72 includes an aspiration needle that can be quickly dried usinga non-contact means, such as forced air or gas, rather than conventionalblotter paper to inhibit blockage of the needle. The flow cell 60 mayinclude chromatography connectors that connect the flow cell 60 totubing or plumbing associated with the flow path 65.

In some embodiments, the analyzer 10 can be configured to direct theaspiration to blow out the injection port immediately after injecting afirst sample before pre-fetching a next sample to maintain liquid-airgaps between neighboring samples.

The sample containers 70 c can be held in beds 71 that can be loaded andplaced in queue for analysis. The samples 70 s are electronicallyassigned a patient identifier to allow electronic correlation to thetest results. Conventionally, the beds 71 include bar codes that areautomatically read and input into the computer as electronic records asa batch of samples, thereby inhibiting adjusting test parameters for aparticular sample. In some embodiments, the NMR analyzer system 10 isconfigured so that the point of identification of each sample is carriedout automatically at the point of aspiration. Thus, an optic or othersuitable reader can be configured to define a patient identifier to aparticular sample while the sample is being aspirated. In any event, thesystem control software 81 can be configured to create an archivablepatient data file record that includes the patient identifier (alsoknown as an accession number) as well as a dilution factor, theNMR-derived measurement values, test date and time, and “common” rackidentifier, where used, and other process information that can beelectronically searched as desired for service, operational and/or auditpurposes. The electronic records can be relayed to a storage location(such as a central collection site within each region or country) and/orstored locally.

In operation, NMR-derived quantitative measurement data for diagnosticclinical reports of patient biosamples can be generated by: (a)automatically serially aspirating biosamples of interest into anenclosed flow path that serially flows the biosamples into an NMRanalyzer having a NMR spectroscopy instrument with a magnet and a boreat a plurality of different clinical sites; (b) automaticallycorrelating a patient identifier to a respective patient biosample; (c)and obtaining NMR derived quantitative measurements of the biosamplesfor diagnostic reports. In some particular embodiments, the operationmay also include (d) automatically monitoring the NMR analyzers at thedifferent clinical sites from a remote system.

Referring again to FIG. 5, the system 10 includes a controller/processor80 that is configured with computer program code 80 s that includesand/or is in communication with instrument automation control software81, analytical software 82, and/or remote communication software 83. Thecontrol software 81 can primarily direct the automated operationalsequences and monitoring protocols of the system 10 while the analyticalsoftware 82 typically includes proprietary software that carries out thequantitative measurements of the biosamples undergoing analysis usingthe NMR-spectrum thereof. For at least the analytical software 82, theprocessor 80 may include a digital signal processor capable ofperforming rapid Fourier transformations.

The remote communication software 83 is configured to carry out and/orfacilitate the communication between the local analyzer(s) and remotecontrol system 10, 15, respectively. The controller/processor 80 may beconfigured as a single processor or a plurality of processors thatcommunicate with each other to provide the desired automated interfacesbetween the system components.

In certain embodiments, it may be desired to maintain the temperature ofthe sample undergoing NMR evaluation at a desired temperature. Forexample, for blood plasma and/or serum samples, it is typically desiredto maintain the temperature of the sample at about 48° C.

In certain embodiments, the system 10 includes a plurality of spatiallydistributed temperature sensors along the flow path 65 that monitor thetemperature of the sample undergoing analysis (not shown). The sampletemperature can be determined at different times in the analysisincluding (a) prior to the sample entering the magnet bore 20 b, (b)prior to initiating the RF pulse sequence, and/or (c) at the time andlocation of discharge from the probe, without disturbing the NMRlineshape in a manner that would impede NMR data collection/reliability.The temperature can be monitored during the NMR data acquisition (suchas at least every 2-5 seconds). The sample can be actively cooled and/orheated during the evaluation to maintain a substantially constanthomogeneous sample temperature without undue thermal gradients.

The system can include cooling and heating means that are configured toprovide distributed heating and/or cooling for reducing hot spots in thesample. One type of heater is a capillary heater that may be slippedover the outer surface of the flow cell 60. An example of a heater isdescribed in U.S. Pat. No. 6,768,304 to Varian, Inc., the contents ofwhich are hereby incorporated by reference herein. It is contemplatedthat a longer capillary heater can be used that extends above the flowcell 60 (where the sample is flowed into the bore 20 b from the top) andmay have a length that is sufficient to extend about a major part of theflow path length. In some embodiments, the system 10 can include aheater that is highly conductive with a relatively large thermal mass(similar to a heat sink) that is above the probe 30 (where the flow isfrom top to bottom), and/or above the flow cell holding portion 60 h tothereby improve distributed heating while reducing the likelihood ofoverheating of the sample as it travels to the probe 30. The largethermal mass may be located outside the magnet bore 20 b.

In some embodiments, a circulating or forced supply oftemperature-controlled gas can be flowed into the magnet bore tomaintain the sample at a desired temperature during the NMR analysis.The temperature of the forced air can be adjusted relatively quickly inresponse to in situ measured sample temperature(s). To reduce moisturethat may be inadvertently directed into sensitive electronics in theprobe or spectrometer, the gas can be filtered and/or dried prior tointroduction into the magnet bore 20 b.

Typically, the samples are preheated from a cooled storage temperature.The auto sample handler 70 can hold the samples while in queue andgradually heat the sample in stages prior to the injection/input port toprovide a sample that is preheated to a desired temperature range (suchas about 45-47.9° C.). Alternatively, the sample may not be heated untilit is in the flow cell 60. In some particular embodiments, the handler70 may also be configured to hold the samples in a refrigerated orcooled state. Combinations of both heating techniques may be used. Thus,the system 10 can include thermal sensors along the path the samplestravel on/in the handler 70 and/or flow path 65 that detect thetemperature thereof and provide real-time feedback to allow the system10 to automatically adjust for any deviation from predicted or norm.

In any event, the system 10 can include a sensor module thatelectronically communicates with processor 80 and accepts/monitorselectronic data output from sensors regarding the status of the sensors.

The flow path 65 may be configured with a valved flow bypass channel(not shown) that bifurcates out of and into the flow path 65 and/or flowcell 60 to allow selected samples to be redirected back into the flowpath 65 above the magnet bore 20 b after the sample exits the probe 30but before it reaches the waste container 61 when a data corruptionevent is detected (not shown). The bypass channel could be in fluidcommunication with a solvent cleaner that allows automatic flushing ofthe bypass channel after use. In other embodiments, the sample(s)affected can be flushed into the waste receptacle and the analyzer 10and/or remote control system 15 can generate a retest notice or orderfor that subject.

Modularity

In certain embodiments, as shown in FIG. 6, the automated clinical NMRanalyzer 10 can be configured with modular assemblies including: anautomated sample handling assembly 70; an NMR spectrometer or detector50 (with a modular NMR probe 30); and a sample flow path 65 with flowcell 60 having a flow cell probe that resides in the NMR spectrometermagnet bore 20 b. Each modular assembly component can be configured toreleasably operate with its mating modular components thereby allowingease of repair and/or field replacement. Further, the analyzer 10 isconfigured with interface software that allows the operationalinterchange between the different modular assemblies. The flow cell 60may be considered a part of the NMR detector 50 or the sample handler 70for modularity purposes. Either way, the NMR analyzer 10 includessuitable interfaces (software and/or hardware) between the automatedsample handler 70 and the NMR detector 50 so as to allow the NMRdetector module 50 and the sample handler module 70 to cooperate toautomatically serially analyze biosamples in a high-volume throughput.

Typically, the NMR analyzer 10 can diagnostically analyze at least about400, and more typically at least about 600, samples per twenty-fourhours. The modular system 10 can be configured so that it can operate ina laboratory environment by staff with little training in NMR supportfunctions. The system 10 may also operate with reduced maintenance anddowntime over conventional NMR detectors and can have a simplified userinterface.

In certain embodiments, the NMR detector 50 can include a flow probe 30that can be modularly replaced in the field and calibrated for operationwithin a relatively short time upon identification of a malfunction orcontamination of the probe 30 due to flow cell leaks and the like.

The system 10 can be configured to store certain operating values of theflow probe 30 being removed and those values can be can bepre-calibrated to defined norms for the new or replacement flow probe30. In particular embodiments, the flow probe 30 can include a memorycard or chip that stores certain operational parameter values (inputupon installation and/or automatically at desired intervals) and can beused in a replacement flow probe 30. In other embodiments, thecapacitors and/or other tunable circuit components can beprogrammatically tuned by an automated tuning routine carried out by theNMR analyzer 10 and/or control system 15.

The flow probe 30 may be configured so that tuning capacitors aremounted underneath (where the flow probe is inserted from the bottominto the bore) or above (where the flow probe is inserted from the topof the magnet bore) the flow probe for easy external access. The flowprobe 30 can be a generally rigid member that is configured toreleasably mount to the magnet without the use of permanent(solder-type) connections.

FIG. 6 illustrates one embodiment of a modular analyzer 10. In thisembodiment, the sample handler assembly 70 includes an upstream portion70 u that provides a staging or queuing sample handler subassembly withautomated drive means and a downstream portion 70 d that includes thesample intake member (such as an injector) 72. Each portion 70 u, 70 dcan have a respective software interface 70I₁, 70I₂ that communicateswith the instrument automation control module 81. The respectiveinterfaces 70I₁, 70I₂ may also optionally communicate with each other.The NMR detector 50 also includes a software interface 50I thatcommunicates with the automation control module 81. The instrumentautomation control module 81 can be configured to interface with acomputer interface and/or network connection circuit/board 50B of theNMR detector 50 (FIG. 7), monitor and/or control sensors, detectors,and/or alarms and direct that certain actions and/or functions becarried out when errors or undue process parameter variations aredetected, provide remote access to the remote station 15 (FIG. 4),directly and/or via the remote communications module 83, and supportautomated start-up and automated (daily) process control monitoring.

As shown in FIG. 6, the instrument automation module 81 optionallycommunicates with the remote communications module 83 a LIS (“LaboratoryInformation System”) interface 84. The LIS interface 84 is incommunication with the LIS system 86 and a user interface module 85 thataccepts local user input into selection of certain operating featuresand/or test report parameters. The LIS interface 84 can be a commoninterface that communicates with other equipment or lab programs,allowing a single common interface that a local user can use in theclinical laboratory. The LIS interface 84 can be in communication withthe analytical software module 82 that includes the test quantificationanalysis or evaluation program code (and may be proprietary and/orcustomized to each type of analysis performed). The data (raw and/or inreport form) can be transmitted to the laboratory's LIS. As indicated bythe broken line connections, the analytical software module 82 mayoptionally communicate directly with either the instrumentationautomation module 81 and/or the remote communications module 83.

The term “module” refers to program code that is directed to carryingout and/or directing particular operational, communications and/ormonitoring functions. The term “module” is not meant to limit theprogram code to a bundled package or a successive portion of code, asthe module program code may be distributed code within a particularprocessor or processors that are in communication. As such, the modulemay be a stand-alone module on a respective single processor or may beconfigured with an architecture/hierarchy that plugs into other programmodules on one or more processors. Furthermore, selected ones or eachmodule noted in the figures may share common code or functionality withother modules.

FIG. 7 illustrates another embodiment of the automated clinical NMRanalyzer 10. As shown, the NMR detector 50 includes an NMR operationalsoftware module 50S with an interface 50I. The NMR software module 50Sis in communication with the remote access/communications module 83. Theremote access/communications module 83 may also be in communication withthe user interface 85. In the embodiment shown, the system 10 includesan electronic library 82L of predetermined computer program functionsthat stores common parameters, or computer program routines, such as aNMR normalization factor, that can be accessed by a plurality ofinterface components so that the common routines or values do not haveto be separately coded in each device/component. As shown, the samplehandler interface 70I, the user interface 85, and the NMR detector 50can access the common library (shown s “dll”) 82L (directly to the NMRdetector as shown and/or optionally via the NMR software module 50S).

The normalization factor is used to standardize the measurements ofdifferent NMR analyzers. Different NMR probes will have different(typically instrument specific) sensitivities based on the “Q” factor ofthe probe. Q is defined as the frequency of the resonant circuit dividedby the half power bandwidth. A standard sample like, for example,trimethyl acetic acid (TMA) can be run on different NMR machines andwith different probes, and the integral of the CH₃ proton can bemeasured to standardize it to a fixed value. The ratio between thepredefined (fixed) value and the integral under then-current conditionsis termed the “normalization factor”, and this can be used tostandardize different NMR analyzers by multiplying any raw NMR intensityby the normalization factor. An extension of the same concept allows foradjusting for relatively small sensitivity differences from day to dayfor the same probe on a particular NMR analyzer by running the samestandard sample and calculating the daily normalization factor in asimilar manner. Hence, the NMR normalization factor can be calculated insitu for each NMR analyzer for each probe and, in some embodiments,adjusted for each NMR analyzer at desired intervals (such as aftercertain numbers of samples, upon start-up, upon detection of a change inselected local operational conditions).

FIG. 8A illustrates yet another embodiment of an exemplary structure ofan NMR analyzer 10. In this embodiment, system coordination software 180communicates with the analytical software 82, the LIS interface 84, theuser interface 85, the NMR control software 50S and the sample handlercontrol/interface software 70I (which can include both the upstream anddownstream interfaces 70I₂, 70I₁, respectively as shown in FIG. 6). Inthis embodiment, hardware components are controlled though the interfacesoftware. The software can provide functionality by exposing acollection of function calls that implement an Application ProgrammingInterface (“API”). The function calls can include mid- and high-levelcommands. For example, in the sample handler 70, “aspirate” or “move tosafe travel height” are mid-level commands. High-level commandsgenerally include multiple mid-level commands which are encompassed by ahigh-level command. For example, a high-level command of “inject sample(x)” implies several mid-level commands be carried out to achieve thisfunction, such as a requirement to move to a safe travel height,position over sample (x), move down, and aspirate sample (x). Examplesof API commands that may be used for certain NMR detector functionsinclude, but are not limited to the following:

-   -   AcquireData ([IN] acquisition parameters, [OUT] fid data) This        command provides the parameter set defining the desired NMR        experiment. The NMR performs the experiment and returns the        acquired data (perhaps an fid) to the calling software.    -   ApplyPhase ([IN] ft data, [IN] phase, [OUT] ft data)    -   AutoPhase ([IN] ft data, [OUT] ft data), [OUT] phase)    -   Calibrate90Pulse ([IN] starting acquisition parameters, [OUT]        ending acquisition parameters)    -   CalibrateTemperatureController( )    -   CenterField ([OUT] field center position)    -   ComputeFt ([IN] processing parameters, [IN] fid data, [OUT] ft        data)    -   GradientShim ([IN] shim map, [OUT] shim values)    -   PhaseLockSignal ([OUT] phase)    -   SetPhase ([IN] phase)    -   SetTemperature ([IN] target temperature)    -   TuneProbe ([IN] channel, [OUT] frequency, [OUT] match value)    -   TuneTemperatureController( )

FIG. 8B is yet another schematic illustration of control and/orcommunication architecture that can be used for the NMR analyzer 10. Asbefore, an instrument automation module 607 can communicate with the NMRdetector 50, the sample handler 70 and the sample intake member 72(which may in some embodiments be an injector). The sample intake member72 may share the sample handler interface 604 and/or be controlledthrough the sample handler 70 in lieu of having its own direct interface605 with the instrument module 607 as shown. In some embodiments, thesample intake member 72 can be configured to aspirate the sample into aflow path 65 as discussed above. In other embodiments, the sample intakemember 72 can be configured to move the sample held in a container intothe NMR detector 50. In any event, as shown, the sample handler 70 andthe NMR detector 50 each include an interface, 604, 606, respectively.

The instrument automation module 607 can communicate with a dataacquisition quality control module 608, a LIS interface 610, aninstrument user interface 617, an NMR Analyzer (“NMRA”) database 611 andan NMRA filebase 612. The system 10 can also include a test automationmodule 613 that allows a selection of different diagnostic test optionsusing the NMR platform. TestB and TestC modules, 615, 616, respectively,can be configured as separate modules that can be deployed as plug inmodules. The test automation module 613 can communicate with the LISinterface 610 the instrument user interface 617, and at leastindirectly, with the instrumentation automation module 607.

Self-Diagnostic/Calibration

FIG. 9 is a flow chart of exemplary operations (blocks 201-230) that canbe executed as a part of an automated self-diagnostic, calibration,and/or tuning start-up procedure that can help assure that the NMRanalyzer 10 is ready for clinical data output before authorizing orallowing evaluation of “real” patient or other target samples. Thestart-up procedure may be self-executing upon operator sign-in orinitiation. The start-up procedure may also be configured to run atdesired intervals, after a certain number of samples are throughput,and/or when the process appears to be out of absolute or relativeprocess limits.

FIG. 10 is a flow chart of exemplary operations (blocks 301-327) thatcan be executed as part of an automated procedure for running qualitycontrol samples through the NMR analyzer 10 including detector 50. Asbefore, the operations can be carried out at start-up and/or at otherdesired intervals. The term “quality control scan” refers to a scantaken of a control reference analyte(s) and/or a biosample to assessoperational status/conditions of the analyzer 10 and/or its environmentat a desired time to assess the operational status or condition of theanalyzer 10 and/or its environment. The reference analyte is configuredto generate a reference peak in an NMR signal. The reference analyte(s)can be provided in a calibration solution of a plurality of differentconstituent chemicals. In some embodiments, the reference analyte isTrimethylacetic acid (“TMA”). In particular embodiments, the TMA is in asolution comprising KCl, CaCl₂, Na₂EDTA and D₂O. However, the referenceanalyte can be any suitable analyte that can generate a reference peakin a NMR signal. In some embodiments, the reference analyte can comprisea molecule that can generate a relatively sharp peak that can be used asa reference for shimming quality and/or to identify the position ofother peaks in the NMR spectrum. Typically, the reference analyte isused qualitatively rather than quantitatively, but may also be usedquantitatively as appropriate.

FIG. 11 is a flow chart of exemplary operations (blocks 501-526) thatcan be executed as part of “normal” operation and/or active-analysis runmode for an automated procedure for running the NMR analyzer 10.

Certain blocks, groups of blocks, and/or combinations of blocks from anyor each of FIGS. 9-11 can be used in particular embodiments.

FIG. 12 is a block diagram of exemplary embodiments of data processingsystems that illustrate systems, methods, and computer program productsin accordance with embodiments of the invention. The Processor 410communicates with the memory 414 via an address/data bus 448. Theprocessor 410 can be any commercially available or custommicroprocessor. The memory 414 is representative of the overallhierarchy of memory devices containing the software and data used toimplement the functionality of the data processing system. 405. Thememory 414 can include, but is not limited to, the following types ofdevices: cache, ROM, PROM, EPROM, EEPROM, flash memory, SRAM, and DRAM.

As shown in FIG. 12, the memory 414 may include several categories ofsoftware and data used in the data processing system 405: the operatingsystem 452; the application programs 454; the input/output (I/O) devicedrivers 458; an automation module 450, which might provide capabilitiessuch as self-adjusting calibration, processing control, or remotecommunications; and data 456.

The data 456 may include NMR signal (constituent and/or compositespectrum lineshape) data 462 which may be obtained from a data or signalacquisition system 420. The data can include values, other operating orprocess parameters of interest, such as leak sensors, thermal sensors,pressure sensors, RF power sensors, the number of successive irregularNMR signal scans, service histories, maintenance files, sample historyfiles, and the like. As will be appreciated by those of skill in theart, the operating system 452 may be any operating system suitable foruse with a data processing system, such as OS/2, AIX or OS/390 fromInternational Business Machines Corporation in Armonk, N.Y., Windows CE,Windows NT, Windows 95, Windows 98, Windows 2000, or Windows XP fromMicrosoft Corporation, Redmond, Wash., Palm OS from PalmSource, Inc.,Sunnyvale, Calif., Mac OS from Apple Computer, Inc, UNIX, FreeBSD, orLinux, proprietary operating systems or dedicated operating systems, forexample, for embedded data processing systems.

The I/O device drivers 458 typically include software routines accessedthrough the operation system 452 by the application programs 454 tocommunicate with devices such as I/O data port(s), data storage 456 andcertain memory 414 components and/or the data acquisition system 420.The application programs 454 are illustrative of the programs thatimplement the various features of the data processing system 405 andpreferably include at least one application that supports operationsaccording to embodiments of the present invention. Finally, the data 454represents the static and dynamic data used by the application programs454, the operating system 452, the I/O device drivers 458, and othersoftware programs that may reside in the memory 414.

While the present invention is illustrative, for example, with referenceto the automation module 450 being an application program in FIG. 12, aswill be appreciated by those of skill in the art, other configurationsmay also be utilized while still benefiting from the teachings of thepresent invention. For example, the automation module 450 may also beincorporated into the operating system 452, the I/O device drivers 458,or other such logical division of the data processing system 405. Thusthe present invention should not be construed as limited to theconfiguration of FIG. 12, which is intended to encompass anyconfiguration capable of carrying out the operations described herein.

In certain embodiments, the automation module 450 may include computerprogram code for communicating with a remote control system (local oroffsite). The automation module 450 can also include program code thatprovides: automated multi-parameter process monitoring andself-correction/adjustment, a log of operational conditions that may becorrelated to patient samples (including time/date data), selectabletest formats and selectable test analysis, a log of data variabilityand/or service history, a log of the number of patient samples processed(which may be parsed over desired intervals), and archived processparameter information for remote interrogation, diagnostics, and otherdata as indicated above.

In particular embodiments, the NMR analyzer 10 can be configured toelectronically monitor (alone and/or cooperating with a remote controlsystem 15) a plurality of components for selected operational variablesand to carry out different testing methodologies according to the testdesired of a particular biosample to facilitate automated function ofthe device automatically whereby the NMR analyzer 10 operates withoutrequiring undue amounts of manual input and/or on-site service supportduring normal operation. Examples of the components and variables werediscussed above and are illustrated in the figures and can include, forexample, one or more of the following:

-   -   electronically monitoring measurements of selected components        and adjusting the operational output/input so that the        component(s) operate within a desired range;    -   electronically automatically calibrating selected electronic        components;    -   executing an automated calibration routine at start-up or other        desired intervals;    -   electronically tuning the flow cell probe;    -   electronically centering a resonance of a sample constituent        (which may be a sample solvent) within an RF window of interest        (i.e., centering a magnetic field in an acquisition window);    -   electronically adjusting lock power and lock phase;    -   electronically shimming the magnet to a desired level of        homogeneity;    -   adjusting the temperature of the flow cell probe;    -   adjusting the temperature of the biosample;    -   electronically calibrating the pulse width of the RF excitation        pulse used to excite the biosample in the magnet bore;    -   electronically (programmatically) determining a normalization        factor to adjust for instrument-specific sensitivity in situ;    -   electronically correlating a biosample with a patient identifier        in situ (such as at the point of aspiration):    -   electronically obtaining the NMR clinical test data from the        biosample and electronically relating the test data to the        patient;    -   electronically controlling the introduction of a reagent(s) to        the biosample prior to obtaining the NMR spectra thereof;    -   electronically controlling the introduction of a selected        calibrant material to the biosample prior to obtaining NMR        spectra;    -   conditioning the biosample to a desired temperature range;    -   obtaining NMR spectra of the biosample using the appropriate NMR        test;    -   obtaining NMR spectra of the biosample and/or a control        validation sample to verify test conditions separate from        obtaining NMR spectra of the biosample for clinical diagnostic        analysis;    -   electronically invalidating, not acquiring, flagging or        discarding NMR spectra for a biosample when test conditions are        outside defined acceptable limits;    -   electronically verifying whether the biosample is delivered        properly to a test location in the magnet bore (such as        confirming the biosample is static or whether it constitutes an        “infinite sample” whereby the sample extends beyond the        detection region so that there are no or reduced boundary        effects);    -   electronically determining whether the delivered biosample has        air bubbles as it resides in the NMR probe flow cell;    -   electronically determining the temperature of the biosample as        it resides in the flow cell (and may include automatically        adjusting the temperature of the biosample in situ if it is        outside acceptable limits);    -   electronically determining whether the suppression of a water        signal is in a desired operational range (and if not        electronically adjusting parameters to adjust the water        suppression to be within the desired range);    -   electronically determining what type of diagnostic test to run        on the biosample under analysis;    -   electronically adjusting experiment protocol parameters based on        the biosample and/or properties thereof;    -   electronically obtaining NMR derived measurements of lipoprotein        particle size(s) and concentrations in a blood plasma and/or        serum sample;    -   electronically determining a patients risk of having and/or        developing CHD and/or Type II diabetes based on NMR derived        lipoprotein measurements;    -   electronically determining an NMR derived diagnostic data        measurement of the biosample and generating an electronic        patient report of the data;    -   electronically obtaining NMR spectra to qualitatively determine        the presence or absence of a selected species or constituent,        subspecies, analyte, interference material, contaminant and/or        toxin; and    -   electronically obtaining NMR spectra to quantitatively determine        the concentration a selected species or constituent, subspecies,        analyte, interference material, contaminant and/or toxin.

The foregoing is illustrative of the present invention and is not to beconstrued as limiting thereof. Although a few exemplary embodiments ofthis invention have been described, those skilled in the art willreadily appreciate that many modifications are possible in the exemplaryembodiments without materially departing from the novel teachings andadvantages of this invention. Accordingly, all such modifications areintended to be included within the scope of this invention as defined inthe claims. In the claims, means-plus-function clauses, where used, areintended to cover the structures described herein as performing therecited function and not only structural equivalents but also equivalentstructures. Therefore, it is to be understood that the foregoing isillustrative of the present invention and is not to be construed aslimited to the specific embodiments disclosed, and that modifications tothe disclosed embodiments, as well as other embodiments, are intended tobe included within the scope of the appended claims. The invention isdefined by the following claims, with equivalents of the claims to beincluded therein.

1. A method of operating a clinical NMR in vitro diagnostic analyzer,comprising: electronically monitoring data associated with a pluralityof selected parameters of the clinical NMR analyzer, the NMR analyzercomprising a flow probe held in a magnet bore of an NMR spectrometer;electronically determining whether the selected parameters are withindesired operational ranges based on the monitored data; initiating anautomated calibration procedure (i) at start-up as part of aself-diagnostic start-up procedure before authorizing or allowingevaluation of patient samples that are introduced to the flow probe heldin the NMR spectrometer and (ii) when one or more of the selectedparameters are determined to be outside desired operational ranges basedon the determining step, wherein the calibration procedure includesdelivering a calibration standard to the flow probe in the NMRspectrometer; automatically aborting a test, alerting an operator ofabnormal conditions and/or adjusting operational parameters of selectedcomponents of the clinical NMR analyzer based on data obtained by theelectronically determining step; introducing patient biosamples to theflow probe in the NMR spectrometer; obtaining NMR signal spectra of theintroduced patient biosamples; and electronically generating at leastone clinical quantitative measurement of the patient biosamples based onthe obtained NMR spectra.
 2. A method according to claim 1, furthercomprising automatically calibrating selected components of the NMRanalyzer based on the monitored data.
 3. A method according to claim 2,wherein the monitoring step monitors a temperature of the probe, andwherein the adjusting step comprises automatically adjusting the probetemperature to within a desired operating range.
 4. A method accordingto claim 3, wherein the automatically adjusting selected operationalparameters comprises tuning the probe to a desired operationalfrequency.
 5. A method according to claim 2, wherein the automaticallyadjusting step comprises automatically electronically centering amagnetic field in an acquisition window.
 6. A method according to claim2, wherein the automatically adjusting step comprises adjusting a lockpower and lock phase associated with the NMR analyzer.
 7. A methodaccording to claim 1, further comprising automatically electronicallyshimming the magnetic field to a desired level of homogeneity based onthe monitored data.
 8. A method according to claim 1, wherein the NMRanalyzer comprises an RF source configured to generate an RF pulse tobiosamples, and wherein the automatically adjusting step comprisesautomatically calibrating the RF source to generate about a 90-degreepulse width within a desired pulse width tolerance.
 9. A methodaccording to claim 2, further comprising automatically determining insitu an appropriate NMR analyzer-specific normalization factor to beused during the analyzing step to thereby adjust for instrument-specificsensitivity.
 10. A method according to claim 1, further comprisingautomatically correlating the obtained NMR data with an identifierassociated with a respective biosample.
 11. A method according to claim10, wherein the correlating comprises electronically relating, at apoint of aspiration, the biosample to an electronic file configured tohold corresponding NMR test data of a respective patient.
 12. A methodaccording to claim 10, further comprising electronically maintaining thecorrelation of the obtained data before, during and after the generatingstep.
 13. A method according to claim 1, further comprisingautomatically adding a desired reagent to a respective biosample priorto the obtaining step; and automatically heating or cooling thebiosample to be within a desired temperature range prior to theobtaining step.
 14. A method according to claim 1, further comprisingelectronically generating a patient retest notice if data corruption iselectronically identified as being associated with a patient test.
 15. Amethod according to claim 1, further comprising adding a referenceanalyte to the biosample prior to the obtaining step.
 16. A methodaccording to claim 1, further comprising verifying test conditions insitu temporally proximate but prior to the obtaining step by obtaining afirst pre-test set of NMR spectra of the biosample or a test validationreference sample and electronically evaluating the first NMR spectratherefrom.
 17. A method according to claim 1, further comprisingelectronically flagging, invalidating and/or discarding the NMR spectraof the biosample when test conditions are identified as being outsidethe acceptable range based on data from the determining step.
 18. Amethod according to claim 1, further comprising a flow cell configuredto hold the biosample in the flow probe in the magnet bore during theobtaining step, wherein the monitoring step comprises determiningwhether a biosample in the NMR flow cell is stationary during theobtaining step.
 19. A method according to claim 1, further comprising anNMR flow cell configured to hold the biosample in the flow probe in themagnet bore during the obtaining step, wherein the determining stepcomprises determining whether a biosample in the NMR flow cell is aninfinite sample during the obtaining step.
 20. A method according toclaim 1, further comprising an NMR flow cell configured to hold thebiosample in the flow probe in the magnet bore during the obtainingstep, wherein the determining step comprises determining whether thereare air bubbles in the biosample residing in the NMR flow cell.
 21. Amethod according to claim 1, further comprising an NMR flow cellconfigured to hold the biosample in the flow probe in the magnet boreduring the obtaining step, wherein the determining step comprisesdetermining a temperature of the biosample in the NMR flow cell, andwherein the automatically adjusting step comprises adjusting parametersthat bring the temperature of the biosample in the NMR flow cell into anacceptable range.
 22. A method according to claim 1, wherein thedetermining and adjusting steps are carried out to perform at least oneof the following: (a) determine whether homogeneity of a magnetic fieldassociated with a magnet in the NMR analyzer is within a desiredoperational range, and automatically shimming the homogeneity if thedetermined homogeneity is outside the desired operational range; and (b)determine whether suppression of a water signal is within a desiredoperational range, and automatically adjusting selected operationalparameters to bring the water suppression to within the desiredoperational range.
 23. A method according to claim 1, wherein thegenerating step automatically electronically generates a plurality ofclinical quantitative measurement of the-biosample, and wherein themethod further comprises automatically generating an electronic patientreport of the measurements.
 24. A method according to claim 1, furthercomprising electronically determining a patient's risk of having and/ordeveloping CHD based on quantitative measurements from the generatingstep.
 25. A method according to claim 1, further comprisingelectronically determining a patient's risk of having and/or developingType II diabetes based on quantitative measurements from the generatingstep.
 26. A method according to claim 1, wherein the generating step iscarried out to electronically obtain NMR derived measurements oflipoprotein particle sizes and concentrations of the biosamples.
 27. Amethod according to claim 1, further comprising electronically obtainingNMR derived measurements of VLDL, LDL and HDL subclass lipoproteinparticles in a blood plasma and/or serum biosample based on thegenerating step.
 28. A method according to claim 1, further comprisingautomatically electronically determining a patient's risk of having aninsulin resistance disorder based on the generating step.
 29. A methodaccording to claim 1, further comprising electronically determining thepresence or absence of a selected species or constituent, subspecies,analyte, interference material, contaminant and/or toxin in thebiosample based on the obtaining and analyzing steps.
 30. A methodaccording to claim 1, further comprising electronically determining theconcentration of a selected species or constituent, subspecies, analyte,interference material, contaminant and/or toxin in the biosample basedon the obtaining and analyzing steps.
 31. A method according to claim 1,further comprising automatically determining a diagnostic test and/orclinical quantitative measurement to be carried out on the biosamplefrom a plurality of different pre-defined tests and/or measurementsusing the NMR analyzer.
 32. A method according to claim 31, furthercomprising adjusting testing parameters based on properties of thebiosample and/or the test to be carried out.
 33. A method according toclaim 1, wherein the initiating step is carried out automatically atequipment startup and/or at intervals during operation.
 34. A methodaccording to claim 1, wherein the introducing, obtaining and generatingsteps are carried out to analyze at least 400 patient biosamples pertwenty-four hours.
 35. A method according to claim 1, wherein theintroducing step is carried out to flowably introduce the patientbiosamples to the NMR spectrometer in a high throughput rate of at leastabout 600 samples per 24 hours.
 36. A method according to claim 34,wherein the introducing, obtaining and generating steps are carried outby the NMR analyzer to diagnostically analyze at least about 400biosamples per twenty-four hours without requiring dedicated on-site NMRsupport staff to reliably operate the NMR analyzer.
 37. A methodaccording to claim 1, wherein the flow probe is a modular top loadingflow probe configuration thereby allowing ease of installation andtuning, the method further comprising releasably attaching the flowprobe by inserting the flow probe into a top of the magnet bore of theNMR spectrometer.
 38. A method according to claim 37, wherein tuningcapacitors associated with the flow probe are mounted above the flowprobe, the method further comprising replacing the flow probe byremoving a first modular flow probe from a top of the magnet, theninserting a second modular flow probe from the top of the magnet andattaching the flow probe to the magnet.
 39. A method according to claim1, wherein the introducing step is carried out using an automated samplehandler with a sample handler interface and a sample injector interface,wherein the clinical NMR analyzer comprises an instrument automationmodule, the method further comprising electronically controlling theoperation of the sample handler, the sample injector and the NMRspectrometer using the instrument automation module.
 40. A methodaccording to claim 39, further comprising electronically selecting whatclinical quantitative measurement to be carried out for a respectivebiosample using the instrument automation module and a patientanalytical test module, wherein the patient analytical test moduleincludes a plurality of selectable different tests.
 41. A methodaccording to claim 39, wherein the instrument automation modulecomprises an LIS interface.
 42. A method according to claim 1, furthercomprising automatically electronically invalidating, not acquiring,flagging or discarding NMR spectra for a respective biosample when testconditions are outside defined limits based on data from the determiningstep.
 43. A method according to claim 1, further comprisingelectronically selecting a test format and/or sample variables ofinterest for a respective biosample and generating a test report for thebiosample using the selected test format.
 44. A method according toclaim 1, wherein the NMR analyzer is configured to generate acustomizable report format, one for each of a plurality of differenttest sites.
 45. A method according to claim 1, wherein the NMR analyzercomprises a sample handler that holds containers of the respectivebiosamples and an enclosed flow path the directs the biosamples to flowinto a flow cell held inside the flow probe in the magnet bore, andwherein the introducing step is carried out by electronically directingan aspirating member to blow out an injection port associated with theflow path after injecting a first sample into the flow path beforepre-fetching a second sample to maintain liquid-air gaps betweenneighboring samples in the flow path.
 46. A method according to claim 1,further comprising: automatically electronically monitor NMR signallineshapes of obtained NMR signal spectra of the biosamples;automatically electronically scale the NMR signal spectra of thebiosamples responsive to the monitored signal lineshape; electronicallycenter a resonance of a biosample constituent within an RF window ofinterest; electronically adjust lock power and lock phase;electronically shim the magnet to a desired level of homogeneity;electronically monitor temperature of the flow cell probe;electronically determine a pulse width of the RF excitation pulse usedto excite the biosample in the magnet bore; electronically monitor anormalization factor to adjust for NMR spectrometer-specific sensitivityin situ; electronically invalidate, not acquire, flag or discard NMRspectra for a biosample when test conditions are determined to beoutside defined acceptable limits; electronically determine what type ofdiagnostic test to run on the biosamples being introduced;electronically obtain NMR derived measurements of lipoprotein particlesize(s) and concentrations in a blood plasma and/or serum sample as thebiosample; electronically relate the test data to respective patients;and electronically generate respective electronic patient reports of themeasurements.
 47. A method according to claim 1, further comprisingautomatically generating a patient data file record for each biosamplethat includes a patient identifier, a dilution factor, the at least oneclinical measurement, and an associated test date and time.
 48. A methodof operating a clinical NMR in vitro diagnostic analyzer, comprising:electronically monitoring data associated with a plurality of selectedparameters of a clinical NMR analyzer; electronically determiningwhether the selected parameters are within desired operational rangesbased on the monitored data, automatically adjusting operationalparameters of selected components of the clinical NMR analyzer based ondata obtained by the electronically determining step; obtaining NMRsignal spectra of a biosample; electronically generating at least oneclinical measurement of the biosample based on the obtained NMR spectra;and before the obtaining step, serially aspirating biosamples fromrespective sample containers into an enclosed flow path andautomatically serially detecting patient identification data for eachrespective patient biosample at a point of aspiration into the enclosedflow path to thereby inhibit incorrect patient biosample correlation.49. A method according to claim 48, further comprising flowing theaspirated samples into a flow cell associated with the flow path held ina magnet bore of the NMR analyzer and directing the sample to travelgenerally vertically to an intermediate location in the magnet boreinside the flow cell held in a flow probe in the magnet bore to therebyreduce flow path length.
 50. A method of operating a clinical NMR invitro analyzer in a patient biosample test laboratory without an onsitededicated NMR support technician, the clinical analyzer comprising anNMR spectrometer with a magnet having a bore, the method comprising thesteps of: automatically introducing in vitro biosamples into atop-loaded flow probe held in the magnet bore of the NMR spectrometer;electronically detecting data associated with selected operatingparameters of the clinical NMR analyzer during normal operation;electronically verifying that selected conditions of the clinical NMRanalyzer are within target operating ranges on the basis of the detecteddata; executing an automated electronic self-diagnostic quality controland/or calibration test during operation using a calibration standardintroduced to the flow probe; and electronically obtaining NMR spectrafor at least 400 biosamples per twenty-four hours and generatingquantitative measurements based on the obtained NMR spectra.